The benefits of gene therapy for alleviating or correcting defects in the production of proteins are widely recognized and a number of gene delivery systems have been developed. One method involves using viruses to deliver the exogenous gene to a cell but this method introduces vital genes along with the new gene and undesirable vital effects may be produced.
Another method involves targeting a polynucleotide to a cell in vivo where it is then expressed. This can be accomplished using a soluble complex of two linked components: 1) a polycation, e.g., poly-L-lysine, that can bind a polynucleotide in a strong but non-damaging interaction and 2) a ligand which can be targeted specifically to a cell surface molecule unique to the cell. See Wu, G. Y. and Wu, C. H. (1988) J. Biol. Chem, 263:14621-14624. The complex specifically binds the targeted cell and is internalized by endocytosis resulting in the incorporation of the polynucleotide.
During endocytosis, the macromolecule to be ingested is enclosed by a small portion of the plasma membrane which pinches off to form an intracellular vesicle or endosome. Newly synthesized lysosomes from the Golgi apparatus are translocated and fused to endosomes. After fusion, the body is called an endolysosome which then develops into a mature lysosome. Lysosomes contain a wide variety of degradative enzymes and, therefore, material in lysosomes can be broken down.
Although transient gene expression may be desirable for some applications of gene therapy, in many other applications, persistent or enhanced expression of the polynucleotide would be required. There exists a need for a method of introducing DNA into cells in vivo so that expression can be enhanced and made to persist. One method for persistent gene expression is to induce replication of the targeted cell(s) (U.S. patent application Ser. No. 588,013, filed Sep. 25, 1990). Zenke et al. recently reported that chloroquine can be used to raise the pH in the targeted cells' lysosomes during "transferrinfection" thus inhibiting degradation of the exogenous gene within lysosomes and increasing the amount of gene incorporated by the cell. See Zenke, M. et al, (May 1990) PNAS USA 87:365-3659. Some investigators have demonstrated that polynucleotide incorporation during transferrinfection can be enhanced by cointernalization of the carrier-polynucleotide complexes with certain viruses. See Curiel et al. (October 1991) PNAS USA 88:8850-8854.